Thyroid cancer is one common endocrine malignancy with various pathological types. MicroRNAs (miRNAs) play essential roles in development, prognosis and treatment of thyroid cancer. However, the role of miR-17-5p in thyroid cancer progression and its mechanism remain poorly understood. The expressions of miR-17-5p and phosphatase and tensin homolog (PTEN) were measured in thyroid cancer tissues and cells by quantitative real-time polymerase chain reaction or western blot. Cell proliferation and apoptosis were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay and flow cytometry, respectively. The protein levels of biomarkers in autophagy or protein kinase B (AKT)/mechanistic target of rapamycin (mTOR) pathway were analyzed by western blot. The interaction between miR-17-5p and PTEN was probed by luciferase activity assay. We found that miR-17-5p expression was elevated and PTEN level was reduced in thyroid cancer tissues and cells compared with their corresponding controls. Knockdown of miR-17-5p or overexpression of PTEN suppressed cell proliferation and autophagy but promoted apoptosis in thyroid cancer cells. PTEN was indicated as a target of miR-17-5p and its interference reversed abrogation of miR-17-5p-mediated inhibition of proliferation and autophagy and increase of apoptosis. Moreover, downregulation of miR-17-5p impeded the activation of AKT/mTOR pathway in thyroid cancer cells, which was attenuated by silencing PTEN. Our data supported that knockdown of miR-17-5p upregulated PTEN expression, therefore leading to suppression of the malignancy of thyroid cancer and inactivation of AKT/mTOR pathway, providing a novel avenue for treatment of thyroid cancer.

#403 Background: Triple negative breast cancer (TNBC: negative for ER, PR and HER2) presents a significant clinical challenge. The frequent occurrence of mutations in TP53, the gene encoding the p53, in TNBC provides an opportunity for therapeutic intervention. p53 is required for cells to respond to DNA damage in the G1 phase of the cell cycle. Tumors with a defective p53 pathway fail to arrest (or undergo apoptosis) in G1 and rely on p53-independent pathways at S and G2 for their survival following DNA damage. Therefore, a potential therapeutic strategy for TP53 mutant tumors is to inhibit the S- and G2-checkpoints. Chk1 is a key regulator of both the S- and G2- checkpoints as such, p53 mutant cells are absolutely dependent on Chk1 activity to respond to DNA damage. This property makes Chk1 a potential therapeutic target in p53 defective tumors. Therefore, we tested the combination of irinotecan (DNA damaging agent) and UCN-01 (non-specific Chk1 inhibitor) in a preclinical model of TNBC. This combination was chosen based on preliminary results obtained in patients with TNBC on a Phase I trial conducted at our institution. UCN-01 is also a potent inhibitor of PDK1 and therefore inhibits the PI3K pathway. Given that PTEN is frequently deficient in TNBC, we also monitored components of the PI3K pathway in our preclinical model of TNBC.
 Material and Method: Tumor biopsies from patients with TNBC were engrafted into the humanized mammary fat pad of immunodeficient NOD/SCID mice. TP53 was sequenced in each engrafted tumor explant and the integrity of the p53 pathway was determined by monitoring p53 stabilization and p21 induction following DNA damage. Three independent TNBC tumor explants, one wild-type and two mutant for TP53 were analyzed for their response to irinotecan and UCN-01 either as single agents or in combination. UCN-01 was administered 24h post irinotecan in mice treated with the combination. Mice were sacrificed 48 h later and tumors were harvested and analyzed for cell cycle arrest (geminin, pCdk1), DNA damage (pChk1, gamma H2AX), apoptosis (cleaved caspase 3), checkpoint bypass (pHistone H3) and the PI3K pathway (pS6) by Western blotting and immunohistochemistry.
 Results: Expression profiling revealed that the characteristics of the tumor are preserved in the tumor explants suggesting this is a valid model system to study experimental therapy for TNBC. Irinotecan induced similar levels of DNA damage in TP53 wild-type and TP53 mutant TNBCs. UCN-01 potently inhibited levels of pS6 independent of p53 status. Strikingly, the combination of irinotecan and UCN-01 selectively induced checkpoint bypass and apoptosis in p53 mutant TNBCs.
 Conclusion: p53 status is a significant predictor of response to combination therapies involving DNA damage followed by Chk1 inhibition. Tumors, like TNBC, that frequently lack a functional p53 pathway may be effectively treated using this strategy. Studies are underway to test different chemotherapy agents and more selective Chk1 and PI3K inhibitors in this preclinical model of TNBC. Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 403.

Background: Women newly diagnosed with primary breast cancer (PBC) often undergo multi-gene panel testing to determine their contralateral breast cancer (BC) risk and whether a contralateral prophylactic mastectomy is warranted. However, with the exception of BRCA1/2, gene-specific associations with contralateral or second PBC (SPBC) have not been established. Methods: The study sample was comprised of 83,278 women with BC referred to a single diagnostic laboratory for multi-gene panel testing. The frequency of pathogenic/likely pathogenic variants in clinically-actionable genes (CAG), including highly penetrant genes (HPG: BRCA1, BRCA2, TP53, PTEN) and moderately penetrant genes (MPG: ATM, CHEK2, PALB2, CDH1, NBN, NF1) was compared between women with a PBC and SPBC. Women with a SPBC <1 year from their first diagnosis were excluded. Logistic regression burden tests were used to test for associations between mutated genes and SPBC adjusting for age at diagnosis of first BC, histology, presence of first- or second-degree relative with BC, and race/ethnicity. Results: The study included 75,550 women with PBC and 7,728 with SPBC. The median (IQR) time between primaries for the SPBC group was 11 (6,17) years . Women with SPBC were slightly more likely to be Caucasian (67.8% vs. 63.4%; p<0.001), older when referred for genetic testing (mean difference 9.7 years, p<0.001), slightly younger at first BC diagnosis (mean difference 2.1 years, p<0.001) and slightly more likely to have >1 first or second degree relative with BC (62.2% vs. 60.8%; p=0.004) than PBC. Among women tested for all CAGs, 4,883 (8.1%) were carriers of pathogenic/likely pathogenic variants (11.1% SPBC vs. 7.8% PBC). CHEK2 was the most frequently mutated gene (3.4% SPBC vs. 2.3% PBC), followed by BRCA1 (2.7% SPBC vs.1.6% PBC), BRCA2 (2.2% SPBC vs. 1.8% PBC), and PALB2 (1.4% SPBC vs. 0.9% PBC). In fully adjusted models, women with SPBC were 1.38 times as likely (p=<0.0001) as women with PBC to test positive for a CAG (OR=1.35 for HPG and 1.34 for MPG). BRCA1 (OR=1.49; p<0.0001), followed by CHEK2 (OR=1.36; p<0.0001) and PALB2 (OR=1.53; p<0.001) were most significantly associated with SPBC. TP53, BARD1, ATM and BRCA2 were marginally associated with SPBC (p=0.01 to 0.06). When results were stratified by race/ethnicity, ORs among Caucasians were similar to those observed overall. Among African Americans, women with SPBC were 1.76 times as likely to carry a CAG (p<0.001) than their PBC counterparts. PALB2 (OR=2.69; p=0.002), BRCA2 (OR=1.85; p=.004), and TP53 (OR=3.88; p=.009) were most significantly associated with SPBC followed by BRCA1 (OR=1.63; p=.002). Analysis of gene associations for other racial/ethnic groups was limited by small sample size. Conclusions: There is a significantly higher prevalence of CAG mutations among women with SPBC, even after adjusting for age at diagnosis and family history. These findings support SPBC as a standalone indication for multigene panel testing. Additional studies aimed to assess cumulative risk of SPBC for CAG beyond BRCA1/2 are needed to help guide clinical management decisions for mutation carriers. Citation Format: Yao K, Clifford J, Li S, LaDuca H, Hulick PJ, Xu J, Gutierrez S, Black MH. Prevalence of genetic mutations in patients with second primary breast cancers [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-09-02.